The effect of glutathione (GSH), GSH transferase, and ascorbic acid on the detoxification of reactive metabolites has been studied. A simple in vitro assay has been devised to study the importance of GSH and GSH transferase on the detoxification of the microsomally generated reactive metabolites of acetaminophen, bromobenzene and naphthalene. These results show that the reactive metabolites of acetaminophen, bromobenzene and naphthalene are more efficiently detoxified in the presence of GSH and GSH transferase than by GSH alone. These data also show that the majority of the reactive epoxide metabolites of bromobenzene and naphthalene are in equilibrium with the aqueous media. These data suggest that the cytochrome P-450 and epoxide hydratase are not a coupled system. Ascorbic acid efficiently detoxified the reactive metabolite of acetaminophen in microsomal incubation mixtures but at doses of 1 gm/kg did not alter the hepatoxicity of acetaminophen. It is concluded that ascorbic acid is not an important detoxification mechanism with acetaminophen.